The major objective of this project is the biochemical identification and characterization of the primary defect in a group of mutants in culture which are pleiotropic for the utilization of alternate carbon sources in place of glucose. In contrast to the parental Chinese hamster lung cell line (V79), these mutants are unable to grow on galactose, fructose, glucose-6-phosphate, glucose-1-phosphate, galactose-1-phosphate, and some do not grow on mannose. The pleiotropic phenotype appears to be caused by a mutation at a single locus. These mutants have been grouped into nine complementation units; six of which are related by an overlapping complementation pattern. Preliminary studies suggest that the pleiotropic carbohydrate mutants are defective in oxidative energy production due to a mutation affecting a citric acid cycle enzyme, component of the electron transport chain, or oxidative phosphorylation. Assays with purified mitochondria reveal that 2 mutants related by the overlapping complementation pattern are defective in the NADH-CoQ reductase portion of the electron transport chain. Two other mutants, in unrelated complementation groups, are defective in both NADH-CoQ reductase and cytochrome oxidase. Another mutant which is temperature-sensitive for growth on galactose may be uncoupled for oxidative phosphorylation at the nonpermissive temperature and coupled at the permissive temperature.